Project Summary Introduction & Hypothesis. The proposed project addresses the role of the human microbiome in the detoxifying arsenic following ingestion. Arsenic poisoning is a significant worldwide threat to public health that leads to a variety of human diseases, including cancer. Polymorphisms in genes involved with arsenic metabolism and transport have been epidemiologically linked to increased risk of lung, skin, bladder, and liver cancer, but there is large inter-individual variability in cancers among similarly exposed individuals, indicating other important factors are involved in disease penetrance. We hypothesize that differences in arsenic metabolism by the gut microbiome, in combination with variability in host metabolism, explains arsenicosis penetrance in exposed populations, and that controlled/engineered arsenic detox in the gut can be used for arsenicosis prevention and treatment. Participants. Co-PI's Walk and McDermott have both led multidisciplinary research projects. Dr. Walk's background in clinical research, germ free mice, and the human microbiome will complement Dr. McDermott's background in arsenic biochemistry and microbial biotransformation. Co-I's Schmidt and Bothner will bring technical expertise regarding advanced murine models, metabolomics, and thiol-targeted proteomics. Collaborators, Drs. X. Chris Le and Samuel Cohen, will bring years of human arsenicosis research experience along with analytical and comparative physiology expertise. Collectively, the assembled team will ensure the successful completion of the proposed research and insightful interpretation of results. What is known? Genes encoding arsenic-active enzymes are present in genomes of human gut microbiome members and gut contents from mouse and humans can metabolize arsenic in vitro. Only three studies have considered the microbiome's role in the production of organo-arsenicals in the host, but no study has experimentally removed the microbiome or established a defined microbiome (gnotobiotic) to test its effects in vivo. Redox and methylation reactions are perhaps the most intensively studied arsenic detoxification mechanisms. However overlapping roles with central cellular metabolism have made manipulation of these pathways difficult and their interactions with the microbiome in arsenic metabolism has not been addressed. What is proposed? --Use germ free mice to model arsenic metabolism in the absence of a microbiome and in gnotobiotic mice mono-associated with engineered E. coli to quantify the influence of specific microbiome arsenical biotransformations on host health. --Study cooperative influences of the microbiome and host redox/methylation in a novel mouse model using metabolomics and thiol-targeted proteomics to uncover arsenical impacts on host metabolism and the proteome. These combined efforts bring novel experimental tools to bear to definitively address detoxification of a prevalent and dangerous human toxin by the human microbiome.